1. Field of the Invention
The present invention relates to a novel constitutive promoter of plant, particularly to a putative lipase promoter Lip3 derived from Arabidopsis, a method for isolating the same, a recombinant expression vector containing the same and uses of the same for improving traits of a farm product. The constitutive promoter of the present invention can be applied to induce a constitutive gene expression such as herbicide resistance gene and reporter gene, because it can express foreign gene regardless of its expression site. In addition, it can be widely used to develop a transgenic plant improving its useful trait.
2. Background of the Related Art
Promoters that are commonly used to express plant genes constitutively or tissue-specifically aiming at transformation should be classified according to their function as follows.
First, there are promoters inducing a constitutive gene expression. One constitutive promoter suitable in a plant may be 35S RNA gene promoter that is derived from cauliflower mosaic virus (CaMV) and being utilized for main promoter of dicotyledons. In addition, actin promoter of rice plant and ubiquitin promoters of corn are commonly used for constitutive promoter suitable in monocotyledons. Recently, cytochrome C (OsOc1) promoter of rice plant is also developed by domestic researchers and being utilized (Korean Patent Registration NO. 10-0429335). These constitutive promoters are mostly introduced into basic vectors for plant transformation so as to promote the expression of antibiotic resistance genes, herbicide resistance genes and reporter genes for selective marker. Therefore, they are considered with first priority to elucidate the function of target gene within a plant in the research aspect.
Second, there are seed-specific promoters. One seed-specific promoter may be a major storage protein promoter of rice plant. In practice, glutelin promoter of rice plant has been used to develop golden rice and being widely applied to induce the seed-specific expression of monocotyledons. In addition, the seed-specific promoters inducing the gene expression of dicotyledons may be lectin promoter derived from peas, napin promoter derived from cabbage, DC-3 promoter derived from carrots and the like. This DC-3 promoter is applied to increasingly produce vitamin E by inducing the gene expression of γ-tocopherol methyl transferase (γ-TMT) in Arabidopsis seeds. Besides, oleosin promoter derived from green perilla is tried to promote the seed-specific expression and already applied for a patent (Korean Patent Application NO. 10-2006-0000783). These seed-specific promoters become a valuable tool to accumulate useful proteins and to produce useful substance from farm plants applicable for seed crops or seed food in itself and raw material of food.
Third, there are specific promoters expressing in roots. The root-specific promoter has not been commercialized yet. However, peroxidase (prxEa) derived from Arabidopsis is separated and identified to be expressed specifically in a root. In addition, several promoters are recently reported to be related with the root-specific expression, after being isolated from mads gene derived from sweet potatoes (ibMADS) and glucose-inducible ADP-glucose pyrophosphatase (AGPase). Furthermore, these promoters are confirmed to promote transient gene expressions root-specifically in carrots and radishes and already registered with patents (Korean Patent Registration NO. 10-0604186 and Korean Patent Registration NO. 10-0604191). Therefore, it is expected that the root-specific promoters should be utilized to accumulate useful proteins and to produce useful substance from root plants applicable for farm crops or food in itself and raw material of food.
Fourth, there are specific promoters for other tissues such as leaves. The tissue-specific promoter may include ribulose bisphosphate carboxylase/oxygenase small subunit (rbcS) promoter that is derived from a rice plant and corn and strongly induces a gene expression exclusively in a photosynthetic tissue such as leaves, RolD promoter that is derived from Agrobacterium and induces a root-specific expression in a plant, patatin promoter that is derived from a potato and induces a tuber-specific expression, phytoene synthase (PDS) promoter that is derived from a tomato and induces a fruit-specific expression in a mature plant, and oleocin promoter that is derived from cabbage and identified to promote a pollen-specific expression in cabbage flowers. The present inventors also have been attempted to induce the gene expression of cytotoxic Bt protein by using a tapetum-specific BcA9 promoter. Then, we have developed a male-sterile plant and registered a patent (Korean Patent Registration No. 10-0435143).
As illustrated above, a lot of promoters have been discovered from various tissues of plants and reported to accomplish their purpose. Presently, they are being investigated and developed industrially. However, most of universal promoters already commercialized or applied by plant researchers may just belong to the above-mentioned cases. Therefore, it is required in the future to actively develop novel promoters controlling gene expressions more accurately according to the developer's intention.
In order to settle above-mentioned problems, the present inventors have tried to exploit novel constitutive promoters of plant. As a consequence, a lipase promoter Lip3 derived from Arabidopsis has been identified to constitutively express an exogenous gene. Then, the Lip3 promoter region was amplified by using a oligonucleotide primer set of SEQ. ID. NO: 6 and SEQ. ID. NO: 7. Furthermore, recombinant expression vectors containing the promoter have been constructed and transformed to Arabidopsis plant. The resulting Arabidopsis has been investigated to measure a degree of constitutive expression with histochemical staining and fluorometric assay of GUS activity, GUS transcripts and the like. Therefore, it is confirmed that the constitutive promoter of the present invention should induce a constitutive expression regardless of its expression site. Hence, the present invention has been completed successfully.